Two industrially important thermophilic bacteria were isolated from Tatapani hot spring located in Kishtwar district of Jammu and Kashmir, India and were named as KSI and KSII. Water sample from Tatapani hot spring was collected for screening of industrially important thermophillic bacteria. Only two thermophilic bacteria (KSI and KSII) were isolated from Tatapani hot spring water sample. The strict thermophilic nature of both the bacterial isolates (KSI and KSII) was tested by growing them at different temperatures ranging from 40°C-70°C. KSI and KSII bacterial isolates showed their optimum growth at temperature 60°C and at pH 9.0. Morphological and biochemical analysis of both the bacterial isolates was carried out. Results revealed that KSI bacterial isolate was non-motile, Gram negative, large rods and whitish in color. While, KSII bacterial isolate was non-motile, Gram positive, medium rods and creamish in color. Molecular identification of both the bacterial isolates (KSI and KSII) was done by 16s rDNA sequencing. Phylogenetic analysis of 16s rDNA sequences of KSI and KSII bacterial isolates obtained after sequencing revealed that KS1 (GenBank accession no KU248487) had closest homology (99%) with Flavobacterium thermophilum G-21 (GenBank accession no NR 104891.1) and bacterial isolate KSII (GenBank accession no KU248486) showed 99% homology with Anoxybacillus sp. DR01 (GenBank accession no EU621359.1). Thermophillic bacteria (KSI and KSII) were tested for production of industrially important thermophilic enzymes. Both KSI and KSII bacterial isolates could produce catalase enzyme (catalase positive), nitrate reductase enzyme (nitrate positive) and could produce acetoin (Voges Proskaue positive). KSI was efficient in producing enzymes such as amylase and cellulase, while KSII was efficient in producing glutaminase. The bacteria (KSI and KSII) isolated in the present study are thermophilic in nature and could produce industrially important thermophilic enzymes. Thus, these microorganisms can be used commercially for large scale production of industrially important thermostable catalase, nitrate reductase, amylase, cellulase and glutaminase enzymes.

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